PROJECT SUMMARY/ABSTRACT: The long-term goal of this project is to better understand and prevent alloimmunization associated with the transfusion of platelets. Alloimmunization to donor MHC antigens following platelet transfusion occurs frequently and can cause complications such as platelet refractoriness and transplant rejection. UV-based pathogen reduction technologies (PRT) were designed to reduce the risk of transfusion-transmission of infectious disease. It has now been shown in mice that PRT has the additional benefit of both preventing alloresponses to treated platelets as well as modulating the response to subsequent untreated alloantigen exposure. The objective of this proposal is to establish a reductionist murine model in order to identify the mechanisms regulating the alloresponse to PRT treated platelets. This includes identifying the antigens required for the response to PRT treated versus untreated allogeneic PRP, identifying the cells that are presenting these antigens, and how this affects the development of a tolerant versus activating alloresponse in the transfusion recipient. The central hypothesis is that indirect presentation of class I MHC alloantigens from apoptotic treated cells drives the immunomodulation observed following transfusion of PRT treated allogeneic PRP; and that this effect is mediated by changes in the localization and activation state of tolerizing DCs, which in turn shifts the T cell response from activating to tolerogenic. The specific aims are: (1) To determine the type and source of alloantigens controlling the response to allogeneic PRP transfusion and the ability of PRT to modulate these responses in vivo; (2) To determine what APC populations are involved in the immune response to untreated and PRT treated allogeneic PRP transfusion in vivo; and (3) To determine the impact of PRT treated and untreated allogeneic PRP transfusion on the activation and differentiation of T cells in vivo. To identify the relevant alloantigens, donor mouse strains will be utilized that are allogeneic only in the MHC region, or only in the class I or class II MHC region, different components of the PRP will be transfused and cell death pathways triggered by PRT will be probed to identify the source of antigen. The relevant APC populations will be determined by measuring the activation, differentiation, and localization of DC subsets in the spleen following transfusion of PRT treated and untreated PRP, and by looking at the role of direct versus indirect presentation. The balance between activating and tolerogenic alloresponses will be assessed by examining the cytokine milieu in vivo and the activation and differentiation of T cells ex vivo following transfusion. Completion of this project will uncover the mechanisms responsible for the immunomodulation observed following PRT treated platelet transfusion and guide efforts to manipulate the immune response to alloantigens for clinical benefit. Increased understanding and control of this response could help improve patient outcomes following allogeneic transfusion, and transplants of solid organ and hematopoietic stem cells.